The specificities and numbers of lymphocytes in animals are tightly controlled to avoid autoimmunity and to avoid accumulation of lymphocytes generated during previous infections. This control can be achieved through the death of autoreactive lymphocytes as consequence of selection events. Similarly, many lymphocytes that are generated in response to infections die when the infectious agent disappears. The lymphocyte repertoire is also influenced by the ability of lymphocytes to alter their antigen receptor. For example, the antigen receptor expressed by some autoreactive lymphocytes can be modified either by deletion of the genes encoding the offending receptor, or by silencing the action of the autoreactive receptor. The numbers, specificities and activities of lymphocytes in animals are controlled, to avoid accumulation of the huge numbers of lymphocytes which are generated in each successive response to infection and to prevent autoimmunity. Several processes are used to achieve this control. Many of the lymphocytes which are generated in response to infections die when the infectious agent disappears. Some autoreactive lymphocytes die, others modify their receptors for antigen, either by deleting genes coding for the offending receptor, or by somehow silencing the action of the autoreactive receptor. Other autoreactive lymphocytes survive for a while as anergic cells and do not respond productively to antigen and have a shortened half life. The Projects in this Program will examine the ways in which lymphocytes are controlled, comparing the mechanisms used in different types of lymphocytes under different circumstances, with the goal of understanding how the immune system generates a useful but innocuous collection of antigen specific lymphocytes. The individual Projects will focus as follows: 1. On the role of bystander, non autoreactive receptors on B cells, and their ability to rescue cells bearing, in addition, autoreactive receptors from death. 2. On the generation and maintenance of anergic B cells, their specificities and the ability of infectious agents to allow these cells to escape anergy. 3. On the role of Bcl-2 related proteins in the death of activated T, and B cells. The Program has 4 Cores, all essential to the work on all Projects. The subjects of the Cores are: Flow Cytometry;Microscopy;Genetics and Administration. PROJECT 1: Mechanisms of B Cell Survival and Death (Pelanda, R.) PROJECT 1 DESCRIPTION (provided by applicant): Millions of B cells are generated daily that express autoreactive antibodies. Recent studies in mice indicate that newly generated immature B cells that react with autoantigens in the bone marrow undergo receptor editing and anergy, and that receptor editing contributes to a significant fraction of the peripheral protective antibody repertoire. These recent studies also suggest that clonal deletion, once thought the predominant mechanism of central tolerance, is probably a default pathway that is carried out when autoreactive B cells are unable to edit their receptors. It is intrinsic to this model that autoreactive B cells must survive for a certain amount of time in order to successfully edit their receptors. Differences in cell survival of autoreactive and non-autoreactive immature B cells ultimately control the selection of these cells and shape the peripheral B cell repertoire. Therefore, we propose that the window of survival of autoreactive immature B cells during which receptor editing takes place is absolutely essential for the development of a protective peripheral antibody repertoire. What physiologically controls the lifespan of immature B cells during receptor editing is not yet clear although likely involves the regulated activity of anti- and pro-apoptotic pathways. In addition to receptor editing, the generation of a specific, effective and innocuous B cell repertoire also relies on the signals that stop further Ig gene rearrangement upon expression of nonautoreactive BCRs and mediate immunoglobulin allelic/isotypic exclusion. Dysfunctions in these pathways may cause either excessive clonal deletion resulting in immunodeficiency or survival of autoreactive B cells resulting in autoimmunity. Available evidences indicate that the expression of non-autoreactive B cell antigen receptors generates tonic signals that are important for survival of non-autoreactive (primary and edited and possibly anergic) immature B cells as well as immunoglobulin allelic/isotypic exclusion. We propose that the NF-?B pathway, BAFF-R signaling and the Bcl-2 family properly translate B cell antigen receptor signaling to regulate cellular lifespan and immunoglobulin gene recombination. We have created mouse strains that generate either nonautoreactive or autoreactive immature B cells, and other strains that have abnormal levels of BCR and BAFF-R expression. Using these biological tools, we propose to determine how the NF-?B pathway and the Bcl-2 family regulate the lifespan of primary immature B cells, and what is the relative contribution of BCR and BAFF-R signaling to cell survival and establishment of Ig allelic/isotypic exclusion. This project will contribute to our understanding of what regulates the lifespan of developing B cells, in particular depending on whether the cells are autoreactive or not. Moreover, our findings will indicate possible mechanisms by which autoreactive B cells escape tolerance and eventually differentiate into autoantibody-forming cells.